Original Article

Comparison of cytokine profile of IFN-γ, IL-5 and IL-10 in cutaneous leishmaniasis using PBMC vs. whole blood

Abstract

Background and Objectives: The surrogate marker (s) of cure and protection in intracellular pathogens is not yet well defined. The aim of this study was to compare the cytokine profile using whole blood cells (WBC) vs. peripheral blood mononuclear cells (PBMC) in healthy and cutaneous leishmaniasis (CL) volunteers.
Materials and Methods: In this study, WBC and PBMC of the volunteers with history of CL (HCL), Active lesion (ACL) and healthy volunteers were collected. The WBC and PBMC were cultured and stimulated with either PHA or soluble Leishmania antigens (SLA), after 72 hours, the supernatants were collected and the levels of IFN-γ, IL-5 and IL-10 were titrated using ELISA method.
Results: The mean ± SD of cytokines using WBC and PBMC in cutaneous leishmaniasis volunteers stimulated with phytohemagglutin (PHA) or SLA are as follow, PHA, IFN-γ=2295±995 vs. 2339±1115, IL-10=853±309 vs. 1330±966, and IL-5=299±136 vs. 352+156, SLA, IFN-γ, 931±824 vs. 825±532, IL-10, 233±78 vs. 408±381, and IL-5, 185±59 vs. 217±76, respectively. There was no significant difference between the IFN-γ, IL-5 and IL-10 levels using WBC vs. PBMC. There was a strong correlation between the cytokine profiles using WBC and PBMC in cutaneous leishmaniasis volunteers.
Conclusion: There was no significant difference between IFN-γ, IL-10, IL-5 levels in whole blood and PBMC of volunteers with active lesion or history of CL. Whole-blood culture which is easier, cheaper and more convenient could be used instead of PBMC to evaluate the cytokine profile in field conditions.

1. Alvar J, Velez ID, Bern C, Herrero M, Desjeux P, Cano J, et al. Leishmaniasis worldwide and global estimates of its incidence. PLoS One 2012;7(5):e35671.
2. Control of the leishmaniases: report of a meeting of the WHO expert commitee on the control of leishmaniases; 2010: Geneva, World Health Organization.
3. DeKrey GK, Lima HC, Titus RG. Analysis of the immune responses of mice to infection with Leishmania braziliensis. Infect Immun 1998;66:827-829.
4. Tripathi P, Singh V, Naik S. Immune response to Leishmania: paradox rather than paradigm. FEMS Immunol Med Microbiol 2007;51:229-242.
5. Sacks D, Noben-Trauth N. The immunology of susceptibility and resistance to Leishmania major in mice. Nat Rev Immunol 2002;2:845-858.
6. Khamesipour A, Dowlati Y, Asilian A, Hashemi-Fesharki R, Javadi A, Noazin S, et al. Leishmanization: use of an old method for evaluation of candidate vaccines against leishmaniasis. Vaccine 2005;23:3642-3648.
7. Saki J, Akhlaghi L, Maraghi S, Meamar AR, Mohebali M. Evaluation of modified Novy-MacNeal-Nicolle medium for isolation of Leishmania parasites from cutaneous lesions of patients in Iran. Res J Parasitol 2009;4:56-62.
8. Scott P, Pearce E, Natovitz P, Sher A. Vaccination against cutaneous leishmaniasis in a murine model. I. Induction of protective immunity with a soluble extract of promastigotes. J Immunol 1987;139:221-227.
9. Mendonca SC, De Luca PM, Mayrink W, Restom TG, Conceicao-Silva F, Da-Cruz AM, et al. Characterization of human T lymphocyte-mediated immune responses induced by a vaccine against American tegumentary leishmaniasis. Am J Trop Med Hyg 1995;53:195-201.
10. Mahmoodi M, Khamesipour A, Dowlati Y, Rafati S, Momeni A, Emamjomeh M, et al. Immune response measured in human volunteers vaccinated with autoclaved Leishmania major vaccine mixed with low dose of BCG. Clin Exp Immunol 2003;134:303-308.
11. Nateghi Rostami M, Seyyedan Jasbi E, Khamesipour A, Mohammadi A. Tumour Necrosis Factor-alpha (TNF-α) and its soluble receptor type 1 (sTNFR I) in human active and healed leishmaniases. Parasite Immunol 2016;38:255-260.
12. Deenadayalan A, Maddineni P, Raja A. Comparison of whole blood and PBMC assays for T-cell functional analysis. BMC Res Notes 2013;6:120.
13. Khamesipour A, Rostami MN, Tasbihi M, Mohammadi AM, Shahrestani T, Sarrafnejad A, et al. Phenotyping of circulating CD8+ T cell subsets in human cutaneous leishmaniasis. Microbes Infect 2012;14:702-711.
14. Habibi GR, Khamesipour A, McMaster W, Mahboudi F. Cytokine gene expression in healing and non-healing cases of cutaneous leishmaniasis in response to in vitro stimulation with recombinant gp63 using semi-quantitative RT–PCR. Scand J Immunol 2001;54:414-420.
15. Rostami MN, Keshavarz H, Edalat R, Sarrafnejad A, Shahrestani T, Mahboudi F, et al. CD8+ T cells as a source of IFN-γ production in human cutaneous leishmaniasis. PLoS Negl Trop Dis 2010;4(10):e845.
16. Nylén S, Khamesipour A, Mohammadi A, Jafari-Shakib R, Eidsmo L, Noazin S, et al. Surrogate markers of immunity to Leishmania major in leishmanin skin test negative individuals from an endemic area re-visited. Vaccine 2006;24:6944-6954.
17. Silva D, Ponte CG, Hacker MA, Antas PR. A whole blood assay as a simple, broad assessment of cytokines and chemokines to evaluate human immune responses to Mycobacterium tuberculosis antigens. Acta Trop 2013;127:75-81.
18. Hodge G, Hodge S, Han P. Increased levels of apoptosis of leukocyte subsets in cultured PBMCs compared to whole blood as shown by Annexin V binding: relevance to cytokine production. Cytokine 2000;12:1763-1768.
19. Deenadayalan A, Heaslip D, Rajendiran AA, Velayudham BV, Frederick S, Yang H-L, et al. Immunoproteomic identification of human T cell antigens of Mycobacterium tuberculosis that differentiate healthy contacts from tuberculosis patients. Mol Cell Proteomics 2010;9:538-549.
20. Damsgaard CT, Lauritzen L, Calder PC, Kjær TM, Frøkiær H. Whole-blood culture is a valid low-cost method to measure monocytic cytokines—a comparison of cytokine production in cultures of human whole-blood, mononuclear cells and monocytes. J Immunol Methods 2009;340:95-101.
21. Mahmoodi M, Rajabalian S, Fekri A, Esfiarpour I. Evaluation of in vitro production of IFN-γ, IL-10, IL-12 and IL-13 by blood cells in patients with cutaneous leishmaniasis lesions. Iran J Allergy Asthma Immunol 2005;4:15-21.
22. Antas PR, Cardoso FL, Oliveira EB, Gomes PK, Cunha KS, Sarno EN, et al. Whole blood assay to access T cell-immune responses to mycobacterium tuberculosis antigens in healthy Brazilian individuals. Mem Inst Oswaldo Cruz 2004;99:53-55.
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IssueVol 11 No 5 (2019) QRcode
SectionOriginal Article(s)
DOI https://doi.org/10.18502/ijm.v11i5.1962
Keywords
Leishmaniasis immune response; Peripheral blood mononuclear cells; Whole blood

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How to Cite
1.
Miramin Mohammadi A, Javadi A, Firooz A, Khamesipour A. Comparison of cytokine profile of IFN-γ, IL-5 and IL-10 in cutaneous leishmaniasis using PBMC vs. whole blood. Iran J Microbiol. 2019;11(5):431-439.